Chymosin is produced by ruminant animals in the lining of the abomasum. Chymosin is produced by gastric chief cells in young ruminants and some other newborn animals to curdle the milk they ingest, allowing a longer residence in the bowels and better absorption. Some other non-ruminant species, including pigs, cats, and seals, produce it.
One study reported finding a chymosin-like enzyme in some human infants, but others have failed to replicate this finding. Humans have a pseudogene for chymosin that does not generate a protein, found on chromosome 1. Humans have other proteins to digest milk, such as pepsin and lipase.:262
Chymosin is used to bring about the extensive precipitation and curd formation in cheese-making. The native substrate of chymosin is K-casein which is specifically cleaved at the peptide bond between amino acid residues 105 and 106, phenylalanine and methionine. The resultant product is calcium phosphocaseinate. When the specific linkage between the hydrophobic (para-casein) and hydrophilic (acidic glycopeptide) groups of casein is broken, the hydrophobic groups unite and form a 3D network that traps the aqueous phase of the milk.
Charge interactions between histidines on the kappa-casein and glutamates and aspartates of chymosin initiate enzyme binding to the substrate. When chymosin is not binding substrate, a beta-hairpin, sometimes referred to as "the flap," can hydrogen bond with the active site, therefore covering it and not allowing further binding of substrate.
Listed below are the ruminant Cym gene and corresponding human pseudogene:
Because of the imperfections and scarcity of microbial and animal rennets, producers sought replacements. With the development of genetic engineering, it became possible to extract rennet-producing genes from animal stomach and insert them into certain bacteria, fungi or yeasts to make them produce chymosin during fermentation. The genetically modified microorganism is killed after fermentation and chymosin is isolated from the fermentation broth, so that the fermentation-produced chymosin (FPC) used by cheese producers does not contain any GM component or ingredient. FPC contains the identical chymosin as the animal source, but produced in a more efficient way. FPC products have been on the market since 1990 and are considered the ideal milk-clotting enzyme.
FPC was the first artificially produced enzyme to be registered and allowed by the US Food and Drug Administration. In 1999, about 60% of US hard cheese was made with FPC and it has up to 80% of the global market share for rennet.